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1.
Proc Natl Acad Sci U S A ; 116(36): 18109-18118, 2019 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-31439814

RESUMO

Preeclampsia (PE) is a dangerous complication of pregnancy, especially when it presents at <34 wk of gestation (PE < 34 wk). It is a major cause of maternal and fetal morbidity and mortality and also increases the risk of cardiometabolic diseases in later life for both mother and offspring. Placental oxidative stress induced by defective placentation sits at the epicenter of the pathophysiology. The placenta is susceptible to activation of the unfolded protein response (UPR), and we hypothesized this may affect mitochondrial function. We first examined mitochondrial respiration before investigating evidence of mitochondrial UPR (UPRmt) in placentas of PE < 34 wk patients. Reduced placental oxidative phosphorylation (OXPHOS) capacity measured in situ was observed despite no change in protein or mRNA levels of electron transport chain complexes. These results were fully recapitulated by subjecting trophoblast cells to repetitive hypoxia-reoxygenation and were associated with activation of a noncanonical UPRmt pathway; the quality-control protease CLPP, central to UPRmt signal transduction, was reduced, while the cochaperone, TID1, was increased. Transcriptional factor ATF5, which regulates expression of key UPRmt genes including HSP60 and GRP75, showed no nuclear translocation. Induction of the UPRmt with methacycline reduced OXPHOS capacity, while silencing CLPP was sufficient to reduce OXPHOS capacity, membrane potential, and promoted mitochondrial fission. CLPP was negatively regulated by the PERK-eIF2α arm of the endoplasmic reticulum UPR pathway, independent of ATF4. Similar changes in the UPRmt pathway were observed in placentas from PE < 34 wk patients. Our results identify UPRmt as a therapeutic target for restoration of placental function in early-onset preeclampsia.


Assuntos
Mitocôndrias/metabolismo , Fosforilação Oxidativa , Pré-Eclâmpsia/metabolismo , Trofoblastos/metabolismo , Resposta a Proteínas não Dobradas , Fatores Ativadores da Transcrição/metabolismo , Chaperonina 60/metabolismo , Fator de Iniciação 2 em Eucariotos/metabolismo , Feminino , Proteínas de Choque Térmico HSP70/metabolismo , Humanos , Mitocôndrias/patologia , Proteínas Mitocondriais/metabolismo , Pré-Eclâmpsia/patologia , Pré-Eclâmpsia/terapia , Gravidez , Trofoblastos/patologia , eIF-2 Quinase/metabolismo
2.
Artigo em Inglês | MEDLINE | ID: mdl-31465879

RESUMO

The ATF/CREB family of transcription factors represents a large group of basic region-leucine zipper (bZip) proteins that regulate diverse cellular responses. Here we carried out a comprehensive analysis of ATF/CREB family members in 22 representative animal species. The family probably originated from the early diverging metazoan and significantly expanded in vertebrates due to multiple whole genome duplication. Duplicates of atf6 were derived from 2R, and duplicates of creb1, crem, jdp2, creb5, atf4, atf5 and atf7 were products of 3R. We also isolated 21 ATF/CREBs, belonging to 6 subfamilies from Nile tilapia. Based on transcriptome data, most members were found to be dominantly expressed in the head kidney, heart, brain and testis. Some ATF/CREBs displayed sexual dimorphic expression in gonad at 5, 90 and 180 dah (days after hatching), but not at 30 dah. creb1a and atf4a were found to be expressed mainly in phase I and II oocytes of the ovary; while creb1b and atf4b mainly in spermatogenic cells of the testis, indicating divergence of duplicated genes from 3R which suggested neofunctionalization or subfunctionalization in gonad. This is the first genome-wide screening and evolutionary analysis of ATF/CREB family in different animals, particularly in teleosts. The expression analysis of this family in tilapia gonad provided a fundamental clue for understanding their important roles in sex differentiation and gonadal development in teleosts.


Assuntos
Fatores Ativadores da Transcrição/metabolismo , Proteína de Ligação a CREB/metabolismo , Ciclídeos/metabolismo , Evolução Molecular , Gônadas/metabolismo , Fatores Ativadores da Transcrição/genética , Animais , Proteína de Ligação a CREB/genética , Ciclídeos/genética , Feminino , Perfilação da Expressão Gênica , Masculino , Ovário/metabolismo , Testículo/metabolismo
3.
Genes Cells ; 24(9): 627-635, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31294895

RESUMO

Cellular senescence plays an important role in aging and is induced by cyclin-dependent kinase (Cdk) inhibitors that accumulate following stresses during aging. However, the underlying mechanism remains elusive. Herein, we demonstrate that activating transcription factor 7 (ATF7), the stress-responsive recruiter of histone H3K9 di- and trimethyltransferases, functions in the accumulation of Cdk inhibitors. Atf7-deficient (Atf7-/- ) mice have a shorter lifespan than wild-type (WT) mice. Levels of p16Ink4a Cdk inhibitor mRNA increased with age more rapidly in Atf7-/- mice than in WT animals. ATF7 binds to the p16Ink4a gene promoter and was released with age. Consistently, histone H3K9me2 levels on the p16Ink4a gene promoter were lower in Atf7-/- mice than in WT animals. Similar results were obtained when Atf7-/- and WT mouse embryonic fibroblasts (MEFs) were cultured under 20% oxygen conditions, which induces cellular senescence via oxidative stress. Phosphorylation of ATF7 by p38 was also increased with the passage of MEFs, consistent with previous observations that ATF7 phosphorylation by p38 induces its release from chromatin. These results indicate that stress-induced and ATF7-dependent epigenetic changes on p16Ink4a genes play an important role in cellular senescence.


Assuntos
Fatores Ativadores da Transcrição/metabolismo , Senescência Celular , Epigênese Genética , Estresse Oxidativo , Fatores Ativadores da Transcrição/genética , Animais , Células Cultivadas , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Feminino , Histonas/genética , Histonas/metabolismo , Longevidade , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Regiões Promotoras Genéticas , Ligação Proteica , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
4.
Am J Physiol Heart Circ Physiol ; 317(2): H472-H478, 2019 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-31274354

RESUMO

The mitochondrial unfolded protein response (UPRmt) is a cytoprotective signaling pathway triggered by mitochondrial dysfunction. UPRmt activation upregulates chaperones, proteases, antioxidants, and glycolysis at the gene level to restore proteostasis and cell energetics. Activating transcription factor 5 (ATF5) is a proposed mediator of the mammalian UPRmt. Herein, we hypothesized pharmacological UPRmt activation may protect against cardiac ischemia-reperfusion (I/R) injury in an ATF5-dependent manner. Accordingly, in vivo administration of the UPRmt inducers oligomycin or doxycycline 6 h before ex vivo I/R injury (perfused heart) was cardioprotective in wild-type but not global Atf5-/- mice. Acute ex vivo UPRmt activation was not cardioprotective, and loss of ATF5 did not impact baseline I/R injury without UPRmt induction. In vivo UPRmt induction significantly upregulated many known UPRmt-linked genes (cardiac quantitative PCR and Western blot analysis), and RNA-Seq revealed an UPRmt-induced ATF5-dependent gene set, which may contribute to cardioprotection. This is the first in vivo proof of a role for ATF5 in the mammalian UPRmt and the first demonstration that UPRmt is a cardioprotective drug target.NEW & NOTEWORTHY Cardioprotection can be induced by drugs that activate the mitochondrial unfolded protein response (UPRmt). UPRmt protection is dependent on activating transcription factor 5 (ATF5). This is the first in vivo evidence for a role of ATF5 in the mammalian UPRmt.


Assuntos
Fatores Ativadores da Transcrição/metabolismo , Doxiciclina/farmacologia , Mitocôndrias Cardíacas/efeitos dos fármacos , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Miócitos Cardíacos/efeitos dos fármacos , Oligomicinas/farmacologia , Resposta a Proteínas não Dobradas/efeitos dos fármacos , Fatores Ativadores da Transcrição/deficiência , Fatores Ativadores da Transcrição/genética , Animais , Modelos Animais de Doenças , Feminino , Regulação da Expressão Gênica , Preparação de Coração Isolado , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias Cardíacas/genética , Mitocôndrias Cardíacas/metabolismo , Mitocôndrias Cardíacas/patologia , Traumatismo por Reperfusão Miocárdica/genética , Traumatismo por Reperfusão Miocárdica/metabolismo , Traumatismo por Reperfusão Miocárdica/patologia , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia
5.
Cell Tissue Res ; 378(3): 427-440, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31309319

RESUMO

The differentiation of sensory neurons involves gene expression changes induced by specific transcription factors. Vomeronasal sensory neurons (VSNs) in the mouse vomeronasal organ (VNO) consist of two major subpopulations of neurons expressing vomeronasal 1 receptor (V1r)/Gαi2 or vomeronasal 2 receptor (V2r)/Gαo, which differentiate from a common neural progenitor. We previously demonstrated that the differentiation and survival of VSNs were inhibited in ATF5 transcription factor-deficient mice (Nakano et al. Cell Tissue Res 363:621-633, 2016). These defects were more prominent in V2r/Gαo-type than in V1r/Gαi2-type VSNs; however, the molecular mechanisms responsible for the differentiation of V2r/Gαo-type VSNs by ATF5 remain unclear. To identify a cofactor involved in ATF5-regulated VSN differentiation, we investigated the expression and function of CCAAT/enhancer-binding protein gamma (C/EBPγ, Cebpg), which is a major C/EBP family member expressed in the mouse VNO and dimerizes with ATF5. The results obtained showed that C/EBPγ mRNAs and proteins were broadly expressed in the postmitotic VSNs of the neonatal VNO, and their expression decreased by the second postnatal week. The C/EBPγ protein was expressed in the nuclei of approximately 70% of VSNs in the neonatal VNO, and 20% of the total VSNs co-expressed C/EBPγ and ATF5 proteins. We examined the trans-acting effects of C/EBPγ and ATF5 on V2r transcription and found that the co-expression of C/EBPγ and ATF5, but not C/EBPγ or ATF5 alone, increased Vmn2r66 promoter reporter activity via the C/EBP:ATF response element (CARE) in Neuro2a cells. These results suggest the role of C/EBPγ on ATF5-regulated VSN differentiation in early postnatal development.


Assuntos
Fatores Ativadores da Transcrição/metabolismo , Proteínas Estimuladoras de Ligação a CCAAT/fisiologia , Células Receptoras Sensoriais , Órgão Vomeronasal , Animais , Diferenciação Celular , Linhagem Celular Tumoral , Camundongos , Camundongos Endogâmicos C57BL , Células Receptoras Sensoriais/citologia , Células Receptoras Sensoriais/metabolismo , Órgão Vomeronasal/crescimento & desenvolvimento , Órgão Vomeronasal/metabolismo
6.
Cancer Sci ; 110(5): 1633-1643, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30891870

RESUMO

Hepatocellular carcinoma (HCC) is a common cancer with poor prognosis. Hepatitis B virus (HBV) is one of the leading causes of HCC, but the precise mechanisms by which this infection promotes cancer development are not fully understood. Recently, miR-340-5p, a microRNA (miRNA) that has been identified as a cancer suppressor gene, was found to inhibit the migration and invasion of liver cancer cells. However, the effect of miR-340-5p on cell proliferation and apoptosis in HBV-associated HCC remains unknown. In our study, we show that miR-340-5p plays an important role during HBV infection and hepatocellular carcinoma development. Specifically, this miRNA directly binds to the mRNA encoding activating transcription factor 7 (ATF7), a protein that both promotes cell proliferation and suppresses apoptosis through its interaction with heat shock protein A member 1B (HSPA1B). We further found that miR-340-5p is downregulated by HBV, which enhances ATF7 expression, leading to enhanced cell proliferation and inhibition of apoptosis. Notably, ATF7 is upregulated in HCC tissue, suggesting that HBV may target miR-340-5p in vivo to promote ATF7/HSPA1B-mediated proliferation and apoptosis and regulate liver cancer progression. This work helps to elucidate the complex interactions between HBV and host miRNAs and further suggests that miR-340-5p may represent a promising candidate for the development of improved therapeutic strategies for HCC.


Assuntos
Fatores Ativadores da Transcrição/genética , Carcinoma Hepatocelular/virologia , Proteínas de Choque Térmico HSP70/genética , Hepatite B/genética , Neoplasias Hepáticas/virologia , MicroRNAs/genética , Fatores Ativadores da Transcrição/metabolismo , Apoptose , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Progressão da Doença , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Proteínas de Choque Térmico HSP70/metabolismo , Células Hep G2 , Hepatite B/metabolismo , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo
7.
PLoS Genet ; 15(2): e1007830, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30789901

RESUMO

The nematode Caenorhabditis elegans has emerged as a genetically tractable animal host in which to study evolutionarily conserved mechanisms of innate immune signaling. We previously showed that the PMK-1 p38 mitogen-activated protein kinase (MAPK) pathway regulates innate immunity of C. elegans through phosphorylation of the CREB/ATF bZIP transcription factor, ATF-7. Here, we have undertaken a genomic analysis of the transcriptional response of C. elegans to infection by Pseudomonas aeruginosa, combining genome-wide expression analysis by RNA-seq with ATF-7 chromatin immunoprecipitation followed by sequencing (ChIP-Seq). We observe that PMK-1-ATF-7 activity regulates a majority of all genes induced by pathogen infection, and observe ATF-7 occupancy in regulatory regions of pathogen-induced genes in a PMK-1-dependent manner. Moreover, functional analysis of a subset of these ATF-7-regulated pathogen-induced target genes supports a direct role for this transcriptional response in host defense. The genome-wide regulation through PMK-1- ATF-7 signaling reveals a striking level of control over the innate immune response to infection through a single transcriptional regulator.


Assuntos
Fatores Ativadores da Transcrição/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/imunologia , Caenorhabditis elegans/microbiologia , Pseudomonas aeruginosa/imunologia , Animais , Caenorhabditis elegans/genética , Imunoprecipitação da Cromatina , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica , Estudo de Associação Genômica Ampla , Imunidade Inata , Sistema de Sinalização das MAP Quinases , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Análise de Sequência de RNA
8.
Anesth Analg ; 129(2): 608-617, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-30303867

RESUMO

BACKGROUND: Propofol can cause degeneration of developing brain cells and subsequent long-term learning or memory impairment. However, at the early stage of embryonic development, the molecular mechanism of propofol-induced inhibition in neural stem cells (NSCs) neurogenesis is still unclear. The aim of this study was to determine the role of propofol in NSCs neurogenesis and, more importantly, to explore the underlying mechanism. METHODS: First, a single intraperitoneal injection of propofol was performed in pregnant mice, and 6 hours after administration of propofol, the hippocampus RNA and the protein of the embryos' brains was extracted to analyze the expression of neuron-specific markers. Second, the primary NSCs were isolated from the hippocampus of mouse embryonic brain and then treated with propofol for cell viability, immunostaining, and transwell assays; more importantly, we performed RNA sequencing (RNA-seq) and q-reverse transcription polymerase chain reaction assays to identify genes regulated by propofol; the Western blot, small interfering RNA (SiRNA), and luciferase reporter assays were used to study the effects of propofol on calmodulin-dependent protein kinase (CaMk) II/5' adenosine monophosphate-activated protein kinase (AMPK)/activating transcription factor 5 (ATF5) signaling pathway. RESULTS: Our results indicated that propofol treatment could inhibit the proliferation, migration, and differentiation of NSCs. The results of RNA-seq assays showed that propofol treatment resulted in downregulation of a group of Ca-dependent genes. The following mechanism studies showed that propofol regulates the proliferation, differentiation, and migration of NSCs through the CaMkII/phosphorylation of serine at amino acid position 485 (pS485)/AMPK/ATF5 signaling pathway. CONCLUSIONS: The results from study demonstrated that propofol inhibits the proliferation, differentiation, and migration of NSCs, and these effects are partially mediated by CaMkII/pS485/AMPK/ATF5 signaling pathway.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Fatores Ativadores da Transcrição/metabolismo , Anestésicos Intravenosos/toxicidade , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Proliferação de Células/efeitos dos fármacos , Hipocampo/efeitos dos fármacos , Células-Tronco Neurais/efeitos dos fármacos , Neurogênese/efeitos dos fármacos , Propofol/toxicidade , Proteínas Quinases Ativadas por AMP/genética , Fatores Ativadores da Transcrição/genética , Animais , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/genética , Movimento Celular/efeitos dos fármacos , Células Cultivadas , Regulação da Expressão Gênica , Hipocampo/enzimologia , Hipocampo/patologia , Camundongos Endogâmicos C57BL , Células-Tronco Neurais/enzimologia , Células-Tronco Neurais/patologia , Transdução de Sinais
9.
Nucleic Acids Res ; 47(1): 283-298, 2019 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-30407559

RESUMO

Various stresses increase disease susceptibility and accelerate aging, and increasing evidence suggests that these effects can be transmitted over generation. Epidemiological studies suggest that stressors experienced by parents affect the longevity of their offspring, possibly by regulating telomere dynamics. Telomeres are elongated by telomerase and shortened by certain stresses as well as telomere repeat-containing RNA (TERRA), a telomere transcript. However, the mechanism underlying the transgenerational effects is poorly understood. Here, we show that TNF-α, which is induced by various psychological stresses, induces the p38-dependent phosphorylation of ATF7, a stress-responsive chromatin regulator, in mouse testicular germ cells. This caused a release of ATF7 from the TERRA gene promoter in the subtelomeric region, which disrupted heterochromatin and induced TERRA. TERRA was transgenerationally transmitted to zygotes via sperm and caused telomere shortening. These results suggest that ATF7 and TERRA play key roles in paternal stress-induced telomere shortening in the offspring.


Assuntos
Fatores Ativadores da Transcrição/genética , Proteínas de Ligação a DNA/genética , Fatores de Transcrição/genética , Transcrição Genética , Fator de Necrose Tumoral alfa/genética , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Animais , Cromatina/genética , Heterocromatina/genética , Humanos , Camundongos , Fosforilação , Regiões Promotoras Genéticas , Estresse Psicológico , Telômero/genética , Encurtamento do Telômero/genética
10.
Sci Rep ; 8(1): 15915, 2018 10 29.
Artigo em Inglês | MEDLINE | ID: mdl-30374102

RESUMO

Synthetic beta-adrenergic agonists (BA) have broad biomedical and agricultural application for increasing lean body mass, yet a poor understanding of the biology underpinning these agents is limiting further drug discovery potential. Growing female pigs (77 ± 7 kg) were administered the BA, Ractopamine (20 ppm in feed), or the recombinant growth hormone (GH), Reporcin (10 mg/48 hrs injected) for 1, 3, 7, 13 (n = 10 per treatment, per time point) or 27 days (n = 15 per treatment). Using RNA-sequencing and inferred pathway analysis, we examined temporal changes to the Longissimus Dorsi skeletal muscle transcriptome (n = 3 per treatment, per time point) relative to a feed-only control cohort. Gene expression changes were affirmed by quantitative-PCR on all samples (n = 164). RNA-sequencing analysis revealed that BA treatment had greater effects than GH, and that asparagine synthetase (Asns) was the 5th most significantly increased gene by BA at day 3. ASNS protein expression was dramatically increased by BA treatment at day 7 (p < 0.05). The most significantly increased gene at day 3 was activating transcription factor 5 (Atf5), a transcription factor known to regulate ASNS gene expression. Gene and protein expression of Atf4, another known regulator of Asns expression, was not changed by BA treatment. Expression of more than 20 known Atf4 target genes were increased by BA treatment, suggesting that BA treatment induces an integrated stress response (ISR) in skeletal muscle of pigs. In support of this, mRNA expression of sestrin-2 (Sesn2) and cyclin-dependant kinase 1 alpha (Cdkn1a), two key stress-responsive genes and negative regulators of cellular growth, were also strongly increased from day 3 of BA treatment. Finally, tRNA charging was the most significantly enriched pathway induced by BA treatment, suggesting alterations to the translational capacity/efficiency of the muscle. BA-mediated changes to the skeletal muscle transcriptome are highly indicative of an integrated stress response (ISR), particularly genes relating to amino acid biosynthesis and protein translational capacity.


Assuntos
Agonistas Adrenérgicos beta/farmacologia , Aspartato-Amônia Ligase/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Músculo Esquelético/metabolismo , Fenetilaminas/farmacologia , Fatores Ativadores da Transcrição/genética , Fatores Ativadores da Transcrição/metabolismo , Aminoacil-tRNA Sintetases/genética , Aminoacil-tRNA Sintetases/metabolismo , Animais , Aspartato-Amônia Ligase/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Feminino , Hormônio do Crescimento/farmacologia , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Suínos
11.
Mol Metab ; 17: 39-48, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30174228

RESUMO

OBJECTIVE: Loss of insulin secretion due to failure or death of the insulin secreting ß cells is the central cause of diabetes. The cellular response to stress (endoplasmic reticulum (ER), oxidative, inflammatory) is essential to sustain normal ß cell function and survival. Pancreatic and duodenal homeobox 1 (PDX1), Activating transcription factor 4 (ATF4), and Activating transcription factor 5 (ATF5) are transcription factors implicated in ß cell survival and susceptibility to stress. Our goal was to determine if a PDX1-ATF transcriptional complex or complexes regulate ß cell survival in response to stress and to identify direct transcriptional targets. METHODS: Pdx1, Atf4 and Atf5 were silenced by viral delivery of gRNAs or shRNAs to Min6 insulinoma cells or primary murine islets. Gene expression was assessed by qPCR, RNAseq analysis, and Western blot analysis. Chromatin enrichment was measured in the Min6 ß cell line and primary isolated mouse islets by ChIPseq and ChIP PCR. Immunoprecipitation was used to assess interactions among transcription factors in Min6 cells and isolated mouse islets. Activation of caspase 3 by immunoblotting or by irreversible binding to a fluorescent inhibitor was taken as an indication of commitment to an apoptotic fate. RESULTS: RNASeq identified a set of PDX1, ATF4 and ATF5 co-regulated genes enriched in stress and apoptosis functions. We further identified stress induced interactions among PDX1, ATF4, and ATF5. PDX1 chromatin occupancy peaks were identified over composite C/EBP-ATF (CARE) motifs of 26 genes; assessment of a subset of these genes revealed co-enrichment for ATF4 and ATF5. PDX1 occupancy over CARE motifs was conserved in the human orthologs of 9 of these genes. Of these, Glutamate Pyruvate Transaminase 2 (Gpt2), Cation transport regulator 1 (Chac1), and Solute Carrier Family 7 Member 1 (Slc7a1) induction by stress was conserved in human islets and abrogated by deficiency of Pdx1, Atf4, and Atf5 in Min6 cells. Deficiency of Gpt2 reduced ß cell susceptibility to stress induced apoptosis in both Min6 cells and primary islets. CONCLUSIONS: Our results identify a novel PDX1 stress inducible complex (es) that regulates expression of stress and apoptosis genes to govern ß cell survival.


Assuntos
Fatores Ativadores da Transcrição/fisiologia , Proteínas de Homeodomínio/fisiologia , Células Secretoras de Insulina/citologia , Transativadores/fisiologia , Fator 4 Ativador da Transcrição/metabolismo , Fator 4 Ativador da Transcrição/fisiologia , Fatores Ativadores da Transcrição/metabolismo , Animais , Apoptose/fisiologia , Linhagem Celular Tumoral , Sobrevivência Celular/fisiologia , Diabetes Mellitus/metabolismo , Modelos Animais de Doenças , Retículo Endoplasmático/fisiologia , Regulação da Expressão Gênica/genética , Genes Homeobox , Proteínas de Homeodomínio/metabolismo , Secreção de Insulina/fisiologia , Células Secretoras de Insulina/metabolismo , Masculino , Camundongos , Estresse Fisiológico/fisiologia , Transativadores/metabolismo , Transcriptoma/genética
12.
Am J Physiol Cell Physiol ; 315(4): C516-C526, 2018 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-29949403

RESUMO

The mitochondrial unfolded protein response (UPRmt) is a protein quality control mechanism that strives to achieve proteostasis in the face of misfolded proteins. Because of the reliance of mitochondria on both the nuclear and mitochondrial genomes, a perturbation of the coordination of these genomes results in a mitonuclear imbalance in which holoenzymes are unable to assume mature stoichiometry and thereby activates the UPRmt. Thus, we sought to perturb this genomic coordination by using a systemic antisense oligonucleotide (in vivo morpholino) targeted to translocase of the inner membrane channel subunit 23 (Tim23), the major channel of the inner membrane. This resulted in a 40% reduction in Tim23 protein content, a 32% decrease in matrix-destined protein import, and a trend to elevate reactive oxygen species (ROS) emission under maximal respiration conditions. This import defect activated the C/EBP homologous protein (CHOP) branch of the UPRmt, as evident from increases in caseinolytic mitochondrial matrix peptidase proteolytic subunit (ClpP) and chaperonin 10 (cpn10) but not the activating transcription factor 5 (ATF5) arm. Thus, in the face of proteotoxic stress, CHOP and ATF5 could be activated independently to regain proteostasis. Our second aim was to investigate the role of proteolytically derived peptides in mediating retrograde signaling. Peptides released from the mitochondrion following basal proteolysis were isolated and incubated with import reactions. Dose- and time-dependent effect of peptides on protein import was observed. Our data suggest that mitochondrial proteolytic byproducts exert an inhibitory effect on protein import, possibly to reduce excessive protein import as a potential negative feedback mechanism. The inhibition of import into the organelle also serves a retrograde function, possibly via ROS emission, to modify nuclear gene expression and ultimately improve folding capacity.


Assuntos
Proteínas de Membrana/genética , Proteínas Mitocondriais/genética , Transdução de Sinais/genética , Resposta a Proteínas não Dobradas/genética , Fatores Ativadores da Transcrição/genética , Animais , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/genética , Transporte Proteico/genética , Proteólise , Espécies Reativas de Oxigênio/metabolismo , Fator de Transcrição CHOP/genética
13.
Mol Cells ; 41(5): 390-400, 2018 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-29754469

RESUMO

Studies have revealed that miR-103a-3p contributes to tumor growth in several human cancers, and high miR-103a-3p expression is associated with poor prognosis in advanced gastric cancer (GC) patients. Moreover, bioinformatics analysis has shown that miR-103a-3p is upregulated in The Cancer Genome Atlas (TCGA) stomach cancer cohort. These results suggest that miR-103a-3p may function as an oncogene in GC. The present study aimed to investigate the role of miR-103a-3p in human GC. miR-103a-3p expression levels were increased in 33 clinical GC specimens compared with adjacent nontumor stomach tissues. Gain- and loss-of-function studies were performed to identify the correlation between miR-103a-3p and tumorigenesis in human GC. Inhibiting miR-103a-3p suppressed GC cell proliferation and blocked the S-G2/M transition in MKN-45/SGC-7901 cells, whereas miR-103a-3p overexpression improved GC cell proliferation and promoted the S-G2/M transition in vitro. Bioinformatics and dual-luciferase reporter assays confirmed that ATF7 is a direct target of miR-103a-3p. Analysis of the TCGA stomach cancer cohort further revealed that miR-103a-3p expression was inversely correlated with ATF7 expression. Notably, silencing ATF7 showed similar cellular and molecular effects as miR-103a-3p overexpression, namely, increased GC cell proliferation, improved CDK2 expression and decreased P27 expression. ATF7 overexpression eliminated the effects of miR-103a-3p expression. These findings indicate that miR-103a-3p promotes the proliferation of GC cell by targeting and suppressing ATF7 in vitro.


Assuntos
Fatores Ativadores da Transcrição/genética , MicroRNAs/fisiologia , Proteínas de Neoplasias/fisiologia , Neoplasias Gástricas/patologia , Regiões 3' não Traduzidas , Fatores Ativadores da Transcrição/antagonistas & inibidores , Adulto , Idoso , Idoso de 80 Anos ou mais , Ciclo Celular/genética , Divisão Celular , Linhagem Celular Tumoral , Estudos de Coortes , Bases de Dados Factuais , Feminino , Mutação com Ganho de Função , Regulação Neoplásica da Expressão Gênica , Genes Reporter , Células HEK293 , Humanos , Mutação com Perda de Função , Masculino , MicroRNAs/genética , Pessoa de Meia-Idade , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , RNA/genética , Neoplasias Gástricas/genética , Transcrição Genética , Transfecção
14.
Nucleic Acids Res ; 46(9): 4487-4504, 2018 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-29490055

RESUMO

Telomeres maintain the integrity of chromosome ends and telomere length is an important marker of aging. The epidemiological studies suggested that many types of stress including psychosocial stress decrease telomere length. However, it remains unknown how various stresses induce telomere shortening. Here, we report that the stress-responsive transcription factor ATF7 mediates TNF-α-induced telomere shortening. ATF7 and telomerase, an enzyme that elongates telomeres, are localized on telomeres via interactions with the Ku complex. In response to TNF-α, which is induced by various stresses including psychological stress, ATF7 was phosphorylated by p38, leading to the release of ATF7 and telomerase from telomeres. Thus, a decrease of ATF7 and telomerase on telomeres in response to stress causes telomere shortening, as observed in ATF7-deficient mice. These findings give credence to the idea that various types of stress might shorten telomere.


Assuntos
Fatores Ativadores da Transcrição/fisiologia , Encurtamento do Telômero , Fator de Necrose Tumoral alfa/fisiologia , Fatores Ativadores da Transcrição/genética , Fatores Ativadores da Transcrição/metabolismo , Animais , Fibroblastos , Células HeLa , Histonas/metabolismo , Humanos , Autoantígeno Ku/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fosforilação , Telomerase/metabolismo , Telômero/metabolismo
15.
J Biol Chem ; 293(8): 2939-2948, 2018 02 23.
Artigo em Inglês | MEDLINE | ID: mdl-29326161

RESUMO

Activating transcription factor 5 (ATF5) is a member of the ATF/cAMP response element-binding protein family of transcription factors. ATF5 regulates stress responses and cell survival, proliferation, and differentiation and also plays a role in viral infections, cancer, diabetes, schizophrenia, and the olfactory system. Moreover, it was found to also have a critical cell cycle-dependent structural function at the centrosome. However, the mechanism that controls the localization of ATF5 at the centrosome is unclear. Here we report that ATF5 is small ubiquitin-like modifier (SUMO) 2/3-modified at a conserved SUMO-targeting consensus site in various types of mammalian cells. We found that SUMOylation of ATF5 is elevated in the G1 phase of the cell cycle and diminished in the G2/M phase. ATF5 SUMOylation disrupted the interaction of ATF5 with several centrosomal proteins and dislodged ATF5 from the centrosome at the end of the M phase. Of note, blockade of ATF5 SUMOylation deregulated the centrosome cycle, impeded ATF5 translocation from the centrosome, and caused genomic instability and G2/M arrest in HeLa cells. Our results indicate that ATF5 SUMOylation is an essential mechanism that regulates ATF5 localization and function at the centrosome.


Assuntos
Fatores Ativadores da Transcrição/metabolismo , Centrossomo/metabolismo , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/metabolismo , Sumoilação , Ubiquitinas/metabolismo , Fatores Ativadores da Transcrição/química , Fatores Ativadores da Transcrição/genética , Sequência de Aminoácidos , Substituição de Aminoácidos , Animais , Linhagem Celular , Centrossomo/enzimologia , Sequência Consenso , Sequência Conservada , Deleção de Genes , Instabilidade Genômica , Proteínas de Fluorescência Verde/química , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Humanos , Microscopia de Fluorescência , Mutagênese Sítio-Dirigida , Mutação , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/metabolismo , Transporte Proteico , Interferência de RNA , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/metabolismo , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/antagonistas & inibidores , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/química , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/genética , Ubiquitinas/antagonistas & inibidores , Ubiquitinas/química , Ubiquitinas/genética
16.
Sci Rep ; 8(1): 857, 2018 01 16.
Artigo em Inglês | MEDLINE | ID: mdl-29339777

RESUMO

The in vivo function of p38 mitogen-activated protein kinase (MAPK) signaling in regulating the response to simulated microgravity is still largely unclear. Using Caenorhabditis elegans as an assay system, we investigated the in vivo function of p38 MAPK signaling in regulating the response of animals to simulated microgravity and the underlying molecular mechanism. Simulated microgravity treatment significantly increased the transcriptional expressions of genes (pmk-1, sek-1, and nsy-1) encoding core p38 MAPK signaling pathway and the expression of phosphorylated PMK-1/p38 MAPK. The pmk-1, sek-1, or nsy-1 mutant was susceptible to adverse effects of simulated microgravity. The intestine-specific activity of PMK-1 was required for its function in regulating the response to simulated microgravity, and the entire p38 MAPK signaling pathway could act in the intestine to regulate the response to simulated microgravity. In the intestine, SKN-1 and ATF-7, two transcriptional factors, were identified as downstream targets for PMK-1 in regulating the response to simulated microgravity. Therefore, the activation of p38 MAPK signaling may mediate a protection mechanism for nematodes against the adverse effects of simulated microgravity. Additionally, our results highlight the potential crucial role of intestinal cells in response to simulated microgravity in nematodes.


Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , MAP Quinase Quinase 4/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Fatores Ativadores da Transcrição/metabolismo , Animais , Proteínas de Caenorhabditis elegans/antagonistas & inibidores , Proteínas de Caenorhabditis elegans/genética , Proteínas de Ligação a DNA/metabolismo , Mucosa Intestinal/metabolismo , MAP Quinase Quinase 4/antagonistas & inibidores , MAP Quinase Quinase 4/genética , Sistema de Sinalização das MAP Quinases , Proteínas Quinases Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases Ativadas por Mitógeno/genética , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/genética , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Análise de Sobrevida , Fatores de Transcrição/metabolismo , Simulação de Ausência de Peso
17.
Cell ; 172(1-2): 218-233.e17, 2018 01 11.
Artigo em Inglês | MEDLINE | ID: mdl-29249357

RESUMO

Signaling pathways that promote adipose tissue thermogenesis are well characterized, but the limiters of energy expenditure are largely unknown. Here, we show that ablation of the anti-inflammatory cytokine IL-10 improves insulin sensitivity, protects against diet-induced obesity, and elicits the browning of white adipose tissue. Mechanistic studies define bone marrow cells as the source of the IL-10 signal and adipocytes as the target cell type mediating these effects. IL-10 receptor alpha is highly enriched in mature adipocytes and is induced in response to differentiation, obesity, and aging. Assay for transposase-accessible chromatin sequencing (ATAC-seq), ChIP-seq, and RNA-seq reveal that IL-10 represses the transcription of thermogenic genes in adipocytes by altering chromatin accessibility and inhibiting ATF and C/EBPß recruitment to key enhancer regions. These findings expand our understanding of the relationship between inflammatory signaling pathways and adipose tissue function and provide insight into the physiological control of thermogenesis that could inform future therapy.


Assuntos
Adipócitos/metabolismo , Montagem e Desmontagem da Cromatina , Metabolismo Energético , Interleucina-10/metabolismo , Termogênese , Fatores Ativadores da Transcrição/metabolismo , Animais , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Linhagem Celular , Células Cultivadas , Interleucina-10/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Transdução de Sinais
18.
Vet Comp Oncol ; 16(1): 102-107, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28480569

RESUMO

BACKGROUND: Activating transcription factor 5 (ATF5) is a transcription factor that is highly expressed in undifferentiated neural progenitor/stem cells as well as a variety of human cancers including gliomas. AIMS: In this study, we examined the expression and localization of ATF5 protein in canine gliomas, and targeting of ATF5 function in canine glioma cell lines. MATERIALS AND METHODS: Paraffin-embedded canine brain glioma tissue sections and western blots of tumours and glioma cells were immunoassayed with anti-ATF5 antibody. Viability of glioma cells was tested with a synthetic cell-penetrating ATF5 peptide (CP-d/n ATF5) ATF5 antagonist. RESULTS: ATF5 protein expression was in the nucleus and cytoplasm and was present in normal adult brain and tumour samples, with significantly higher expression in tumours as shown by western immunoblotting. CP-d/n ATF5 was found to decrease cell viability in canine glioma cell lines in vitro in a dose-dependent manner. CONCLUSION: Similarities in expression of ATF5 in rodent, dog and human tumours, and cross species efficacy of the CP-d/n ATF5 peptide support the development of this ATF5-targeting approach as a novel and translational therapy in dog gliomas.


Assuntos
Fatores Ativadores da Transcrição/metabolismo , Neoplasias Encefálicas/veterinária , Doenças do Cão/metabolismo , Glioma/veterinária , Fatores Ativadores da Transcrição/imunologia , Animais , Anticorpos Antineoplásicos/imunologia , Western Blotting/veterinária , Neoplasias Encefálicas/imunologia , Neoplasias Encefálicas/metabolismo , Linhagem Celular Tumoral , Cães , Glioma/imunologia , Glioma/metabolismo
19.
Sci Rep ; 7(1): 16675, 2017 11 30.
Artigo em Inglês | MEDLINE | ID: mdl-29192290

RESUMO

Recently, it has been reported that human hepatocyte-like cells can be generated from fibroblasts by direct reprogramming technology. However, the conversion efficiency of human induced hepatocyte-like cells (hiHeps) is not high enough. In addition, comparative analysis with the existing models of hepatocytes, such as human iPS cell-derived hepatocyte-like cells and primary human hepatocytes, has not been sufficiently carried out. In this study, we screened hepatic transcription factors for efficient direct hepatic reprogramming and compared hepatic functions between hiHeps and other existing hepatocyte models. We found that human fibroblasts were efficiently converted into hiHeps by using a combination of ATF5, PROX1, FOXA2, FOXA3, and HNF4A (albumin+/alpha-1 antitrypsin+ cells = 27%, asialoglycoprotein receptor 1+ cells = 22%). The CYP expression levels and CYP activities in hiHeps were higher than those in human iPS cell-derived hepatocyte-like cells, but lower than those in short-term (4 hr) cultured primary human hepatocytes and primary human hepatocytes collected immediately after thawing. These results suggested that functional hiHeps could be efficiently generated by ATF5, PROX1, FOXA2, FOXA3, and HNF4A transduction. We believe that hiHeps generated by our method will be useful for the drug-discovery activities such as hepatotoxicity screening and drug metabolism tests.


Assuntos
Transdiferenciação Celular/genética , Reprogramação Celular/genética , Fibroblastos/citologia , Fibroblastos/metabolismo , Hepatócitos/citologia , Hepatócitos/metabolismo , Fatores de Transcrição/genética , Fatores Ativadores da Transcrição/genética , Fatores Ativadores da Transcrição/metabolismo , Biomarcadores , Linhagem Celular , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Fator 3-beta Nuclear de Hepatócito/genética , Fator 3-beta Nuclear de Hepatócito/metabolismo , Fator 3-gama Nuclear de Hepatócito/genética , Fator 3-gama Nuclear de Hepatócito/metabolismo , Fator 4 Nuclear de Hepatócito/genética , Fator 4 Nuclear de Hepatócito/metabolismo , Humanos , Fatores de Transcrição/metabolismo , Transcriptoma , Transdução Genética , Transgenes
20.
Acta Neuropathol ; 134(6): 839-850, 2017 12.
Artigo em Inglês | MEDLINE | ID: mdl-28861715

RESUMO

Activating transcription factor-5 (ATF5) is a stress-response transcription factor induced upon different cell stressors like fasting, amino-acid limitation, cadmium or arsenite. ATF5 is also induced, and promotes transcription of anti-apoptotic target genes like MCL1, during the unfolded protein response (UPR) triggered by endoplasmic reticulum stress. In the brain, high ATF5 levels are found in gliomas and also in neural progenitor cells, which need to decrease their ATF5 levels for differentiation into mature neurons or glia. This initially led to believe that ATF5 is not expressed in adult neurons. More recently, we reported basal neuronal ATF5 expression in adult mouse brain and its neuroprotective induction during UPR in a mouse model of status epilepticus. Here we aimed to explore whether ATF5 is also expressed by neurons in human brain both in basal conditions and in Huntington's disease (HD), where UPR has been described to be partially impaired due to defective ATF6 processing. Apart from confirming that ATF5 is present in human adult neurons, here we report accumulation of ATF5 within the characteristic polyglutamine-containing neuronal nuclear inclusions in brains of HD patients and mice. This correlates with decreased levels of soluble ATF5 and of its antiapoptotic target MCL1. We then confirmed the deleterious effect of ATF5 deficiency in a Caenorhabditis elegans model of polyglutamine-induced toxicity. Finally, ATF5 overexpression attenuated polyglutamine-induced apoptosis in a cell model of HD. These results reflect that decreased ATF5 in HD-probably secondary to sequestration into inclusions-renders neurons more vulnerable to mutant huntingtin-induced apoptosis and that ATF5-increasing interventions might have therapeutic potential for HD.


Assuntos
Fatores Ativadores da Transcrição/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Doença de Huntington/metabolismo , Corpos de Inclusão/metabolismo , Neurônios/metabolismo , Peptídeos/metabolismo , Animais , Apoptose , Caenorhabditis elegans , Linhagem Celular Tumoral , Modelos Animais de Doenças , Estresse do Retículo Endoplasmático/fisiologia , Humanos , Proteína Huntingtina/genética , Proteína Huntingtina/metabolismo , Doença de Huntington/patologia , Corpos de Inclusão/patologia , Camundongos Transgênicos , Neurônios/patologia , Neuroproteção/fisiologia
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